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. 2017 May 18;7:2075. doi: 10.1038/s41598-017-02129-w

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Mitochondrial Kv3.4 was more sensitive to CoCl₂ and PX-478 (A) Immunocytochemical analysis demonstrated that Kv3.4 co-localizes with mitochondria, and co-localized Kv3.4 decreased after 6 h of 100 µM CoCl₂ treatment (green: Kv3.4, red: mitochondria, yellow: co-localization). We assessed five different images in each group (the control and CoCl₂ treatment groups) to avoid cherry-picking; representative images are shown. (B) Mitochondrial fractionation demonstrated that Kv3.4 is present in both the mitochondria and cytosol and that mitochondrial 100 kDa Kv3.4 is sensitive to CoCl₂ alone or CoCl₂ with PX-478 treatment. Kv3.4 (100 kDa) was down-regulated by 4 h of CoCl₂ treatment, whereas 4 h of PX-478 pretreatment followed by 4 h of CoCl₂ and PX-478 treatment induced the accumulation of Kv3.4. COX-4 and α-tubulin are used as mitochondrial and cytosolic markers, respectively. All experiments were performed in quadruplicate; representative images are shown.